Limulus polyphemus Hemocyanin: 10 Å Cryo-EM Structure, Sequence Analysis, Molecular Modelling and Rigid-body Fitting Reveal the Interfaces Between the Eight Hexamers

Abstract The blue copper protein hemocyanin from the horseshoe crab Limulus polyphemus is among the largest respiratory proteins found in nature (3.5 MDa) and exhibits a highly cooperative oxygen binding. Its 48 subunits are arranged as eight hexamers (1×6mers) that form the native 8×6mer in a nested hierarchy of 2×6mers and 4×6mers. This quaternary structure is established by eight subunit types (termed I, IIA, II, IIIA, IIIB, IV, V, and VI), of which only type II has been sequenced. Crystal structures of the 1×6mer are available, but for the 8×6mer only a 40 A 3D reconstruction exists. Consequently, the structural parameters of the 8×6mer are not firmly established, and the molecular inte…